Chromatin immunoprecipitation of Acetyl-Histone H3 (Lys9/Lys14) was performed on cross-linked chromatin from 1 x 10^6 HTC-IR rat hepatoma cells treated with insulin for 0, 5, or 20 minutes. Immunoprecipitation was performed using a multiplex microplate Matrix ChIP assay (see reference for Matrix ChIP protocol: http://www.ncbi.nlm.nih.gov/pubmed/22098709) with 1ul/100ul well volume of an Acetyl-Histone H3 (Lys9/Lys14) polyclonal antibody (Product # PA5-16194). Chromatin aliquots from ~1 x 10^5 cells were used per ChIP pull-down. Quantitative PCR data were done in quadruplicate using 1ul of eluted DNA in 2ul SYBR real-time PCR reactions containing primers to amplify 7kb upstream of the Egr1 gene, promoter-exon1 (transcription start site, TSS) of Egr1, or exon-2 of Egr1. PCR calibration curves were generated for each primer pair from a dilution series of sheared total rat genomic DNA. Quantitation of immunoprecipitated chromatin is presented as signal relative to the total amount of input chromatin. Results represent the mean +/- SEM for three experiments. A schematic representation of the Egr-1 locus is shown above the data where boxes represent exons (black boxes = translated regions, white boxes = untranslated regions), the zigzag line represents an intron, and the straight line represents upstream sequence. Regions amplified by Egr-1 primers are represented by the black bars above the schematic. Data courtesy of the Innovators Program.
|Tested species reactivity||C. elegans, Human, Mouse, Non-human primate, Rat, Zebrafish|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to the amino terminus of histone H3 acetylated on lysines 9 and 14|
|Purification||Antigen affinity chromatography|
|Storage buffer||0.01M HEPES, pH 7.5, with 0.15M NaCl, 100µg/ml BSA, 50% glycerol|
|Tested Applications||Dilution *|
|ChIP assay (ChIP)||1:50|
|Immunohistochemistry (Paraffin) (IHC (P))||1:800|
|Western Blot (WB)||1:1000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
It is not recommended to aliquot this antibody.
This antibody is not cross-reactive with other acetylated histones.
Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Two molecules of each of the four core histones (H2A, H2B, H3, and H4) form an octamer, around which approximately 146 bp of DNA is wrapped in repeating units, called nucleosomes. The linker histone, H1, interacts with linker DNA between nucleosomes and functions in the compaction of chromatin into higher order structures. This gene is intronless and encodes a member of the histone H3 family. Transcripts from this gene lack polyA tails but instead contain a palindromic termination element. This gene is found in the large histone gene cluster on chromosome 6.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.